Parathyroid hormone (PTH) is a major regulator of mineral homeostasis and bone metabolism. Intermittent PTH therapy is approved for treatment of osteoporosis, yet the cellular mechanisms underlying the biologic effects of PTH are not completely understood, including the molecular basis for the observation that low-dose, intermittent administration of PTH elicits net bone formation, whereas continuous administration of high-dose PTH causes predominantly bone loss. The biological actions of PTH are mediated by a G protein-coupled receptor (PTH1R). We demonstrated that a cytoplasmic molecule, Beta-arrestin2 (beta-arr2), regulates the activity of PTH1R and its agonists in vitro by promoting rapid endocytosis of ligand--receptor complexes and by inhibiting cAMP signaling in response to agonists In this revised application, we present preliminary showing that compared to wild-type mice, beta-arr2 null mice have reduced bone mass and an altered bone response following intermittent PTH administration. In addition, osteoblasts from mice null for beta-arr2 exhibit increased and sustained cAMP signaling in response to PTH. We therefore hypothesize that fl-arrestin2 is a key modulator of the biologic activity of PTH in bone. We will pursue three specific aims to test this hypothesis: 1) Determine the role of beta-arr2 in regulating the anabolic response of bone to intermittent PTH, by determining the response of ovariectomized beta-arr2 KO and WT mice to intermittent PTH therapy. 2) Determine the cellular and molecular mechanisms of PTH biologic activity in primary osteoblastic cell cultures from beta arrestin2 KO mice. 3) Determine the effects of targeted overexpression of beta-arrestin2 in osteoblasts on the skeletal response to intermittent and continuous PTH administration in ovariectomized and intact mice, respectively. In summary, the overall goal of this project is to improve our understanding of the mechanisms regulating the biologic activity of PTH in bone. By conducting complementary in vivo, ex vivo, and in vitro experiments in mice deficient for beta-arrestin2 and in mice overexpressing beta- arrestin-2 specifically in bone, we will provide novel insights into potential mechanisms that underlie the distinct skeletal response to intermittent versus continuous PTH administration. Information gained from the proposed studies will be instrumental for the development of new PTH1R ligands with improved signaling and biologic activity profiles for treatment of osteoporosis and other metabolic bone disorders.